1887

Abstract

Summary: adherence to epithelial cells is the first step in the infectious process, but in spite of its importance, current methods for the quantitative measurement of adherence of to epithelial cells have some serious limitations. They are based on filtration assays and either microscopic or radiometric analysis. The adherence reaction is usually carried out with a large excess of yeasts (100-fold) over epithelial cells in order to perform the microscopic analysis, which is slow, subjective and limited to 100-200 cells and thus lacks statistical power. The radiometric analysis fails to measure individual cells. A method for measuring yeast adherence that overcomes these problems has been developed. It is based on labelling the yeasts with the fluorogenic marker 2′,7′-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester (BCECF) prior to the adherence reaction, and analysing 10 epithelial cells by flow cytometry, while nonbound yeasts are excluded by gating. Two subpopulations of buccal epithelial cells (BECs) which differ in their mean fluorescence intensities per cell (MFIs) were observed: one with MFI which did not exceed nonspecific fluorescence, and the other with MFI as high or higher than the MFI of labelled yeasts. The two subpopulations represent yeast-free and yeast-binding epithelial cells, respectively, and the MFI increment of the BECs is a quantitative measure of the extent of yeast adherence. Control experiments confirming previously described basic features of adherence, such as enhanced adherence at increasing yeast excess, diminished adherence of trypsin-treated or heat-inactivated yeasts, and the differential adherence of various species, supported the validity of the assay. The possibility of studying adherence reliably at low yeast: epithelial cell ratios, which better mimic adhesion as it occurs , is an important advantage of the assay. New findings, using this method, included the observation that exfoliated BECs from diabetic patients exhibited the same capacity for adherence as cells from healthy controls, and that epithelial cells from early human ontogenic stages had a significantly lower adherence level than those from later stages.

Loading

Article metrics loading...

/content/journal/micro/10.1099/13500872-141-7-1523
1995-07-01
2024-05-08
Loading full text...

Full text loading...

/deliver/fulltext/micro/141/7/mic-141-7-1523.html?itemId=/content/journal/micro/10.1099/13500872-141-7-1523&mimeType=html&fmt=ahah

References

  1. Calderone R.A. 1993; Recognition between Candida albicans and host cells.. Trends Microbiol 1:55–58
    [Google Scholar]
  2. Calderone R.A., Braun P.C. 1991; Adherence and receptor relationships of Candida albicans. . Microbiol Rev 55:1–20
    [Google Scholar]
  3. Cox F. 1986; Candida albicans adherence in newborn infants.. J Med Vet Mycol 24:121–125
    [Google Scholar]
  4. Cutler J.E. 1991; Putative virulence factors of Candida albicans. . Anna Rev Microbiol 45:187–218
    [Google Scholar]
  5. Darwazeh A.M., Lamey P.J., Samaranayake L.P., MacFarlane T.W., Fisher B.M., Macrury S.M., MacCuish A.C. 1990; The relationship between colonisation, secretor status and in vitro adhesion of Candida albicans to buccal epithelial cells from diabetics.. J Med Microbiol 33:43–49
    [Google Scholar]
  6. Edwards J.E.J., Gaither T.A., O’Shea J.J., Rotrosen D., Lawley T.J., Wright S.A., Frank M.M., Green I. 1986; Expression of specific binding sites on Candida with functional and antigenic characteristics of human complement receptors.. J Immunol 137:3577–3583
    [Google Scholar]
  7. Goldenhersh M.A., Good R.A., Sarkar N.H., Safai B. 1982; Separation of epidermal cells by density gradient centrifugation on a continuous colloidal silica (Percoll) gradient.. Anal Biochem 119:246–252
    [Google Scholar]
  8. Haughland R.P. 1992 Handbook of Fluorescent Probes and Research Chemicals 1992-1994, 5th edn.. p. 112 Eugene, OR: Molecular Probes:
    [Google Scholar]
  9. Hostetter M.K. 1994; Adhesins and ligands involved in the interaction of Candida spp. with epithelial and endothelial surfaces.. Clin Microbiol Rev 7:29–42
    [Google Scholar]
  10. Huisjes H.J. 1973; Cytology of the amniotic fluid and its clinical applications . In Amniotic Fluid Research and Clinical Application pp. 95–132 Fairweather D.V.I., Eskes T.K.A.B. Edited by Amsterdam: Excerpta Medica;
    [Google Scholar]
  11. Kennedy M.J. 1988; Adhesion and association mechanisms of Candida albicans. . Curr Top Med Mycol 2:73–169
    [Google Scholar]
  12. Kennedy M.J. 1990; Models for studying the role of fungal attachment in colonization and pathogenesis.. Mycopathologia 109:123–137
    [Google Scholar]
  13. Kennedy M.J., Calderone R.A., Cutler J.E., Kanabe T., Riesselman M.H., Robert R., Senet J.M., Annaix V., Bouali A., Mahaza C, Bouchara J.-P., Miegeville M., Marot-Leblond A., Segal E. 1992; Molecular basis of Candida albicans adhesion.. J Med Vet Mycol 30:95–122
    [Google Scholar]
  14. Kimura LH., Pearsall N.N. 1978; Adherence of Candida albicans to human buccal epithelial cells.. Infect Immun 21:64–68
    [Google Scholar]
  15. King R.D., Lee J.C., Morris A.L. 1980; Adherence of Candida albicans and other Candida species to mucosal epithelial cells.. Infect Immun 27:667–674
    [Google Scholar]
  16. Klotz S.A., Drutz D.J., Zajic J.E. 1985; Factors governing adherence of Candida species to plastic surfaces.. Infect Immun 50:97–101
    [Google Scholar]
  17. Kozinn P.J., Taschdjian C.L., Wiener H. 1958; Incidence and pathogenesis of neonatal candidiasis.. Pediatrics 21:421–429
    [Google Scholar]
  18. Kwon-Chung K. J., Bennett J. E. 1992; Candidiasis.. In Medical MycologyMycology pp. 280–336 Philadelphia & London: Lea & Febiger;
    [Google Scholar]
  19. Lamey P.J., Darwaza A., Fisher B.M., Samaranayake L.P., MacFarlane T.W. 1988; Secretor status, candidal carriage and candidal infection in patients with diabetes mellitus.. J Oral Pathol 17:354–357
    [Google Scholar]
  20. Martin E., Bhakdi S. 1991; Quantitative analysis of opsono- phagocytosis and of killing of Candida albicans by human peripheral blood leukocytes by using flow cytometry.. J Clin Microbiol 29:2013–2023
    [Google Scholar]
  21. Odds F.C. 1988 Candida and Candidosis, 2nd edn.. London: Bailliére Tindall;
    [Google Scholar]
  22. Odds F. C. 1994; Candida species and virulence.. ASM News 60:313–318
    [Google Scholar]
  23. Rotrosen D., Calderone R.A., Edwards J.E.J. 1986; Adherence of Candida species to host tissues and plastic surfaces.. Rev Infect Dis 8:73–85
    [Google Scholar]
  24. Segal E. 1994; Virulence factors . In Pathogenic Yeasts and Yeast Infections pp. 49–60 Segal E., Baum G.L. Edited by Boca Raton, FL: CRC Press;
    [Google Scholar]
  25. Segal E., Soroka A., Schechter A. 1984; Correlative relationship between adherence of Candida albicans to human vaginal epithelial cells in vitro and candidal vaginitis.. Sabouraudia 22:191–200
    [Google Scholar]
  26. Shapiro H.M. 1988 Practical Flow Cytometry, 2nd edn.. New York: A.R.Liss;
    [Google Scholar]
  27. Siegel S., Castellan N.J. 1988 Non-parametric Statistics for the Behavioral Sciences pp. 80–87 3rd edn. New York: McGraw-Hill;
    [Google Scholar]
  28. Sobel J.D. 1993; Candidal vulvovaginitis.. Clin Obstet Gynecol 36:153–165
    [Google Scholar]
  29. Sobel J.D., Myers P.G., Kaye D., Levison M.E. 1981; Adherence of Candida albicans to human vaginal and buccal epithelial cells.. J Infect Dis 143:76–82
    [Google Scholar]
  30. Srebrnik A., Segal E. 1990; Comparison of Candida albicans adherence to human corneocytes from various populations.. Acta Dermato-Venereol(Stockh) 70:459–462
    [Google Scholar]
  31. Yeaman M.R., Sullam P.M., Dazin P.F., Ghannoum M.A., Edwards J.E., Bayer A.S. 1994; Fluconazole and platelet microbicidal protein inhibit Candida adherence to platelets in vitro.. Antimicrob Agents Chemother 38:1460–1465
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/13500872-141-7-1523
Loading
/content/journal/micro/10.1099/13500872-141-7-1523
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error